Nitazoxanide (NTZ; 2-(acetyloxy)-N-(5-nitro-2-thiazolyl)benzamide) is an FDA approved antiparasitic drug useful for the treatment of Giardia and Cryptosporidium infections in adults and children. It is a broad spectrum antiparasitic and anti-diarrheal. NTZ efficacy is limited by its poor solubility and strong binding and inactivation by serum proteins.
Coagulase-negative staphylococci (CoNS) have emerged as important opportunistic hospital acquired pathogens that are the leading cause of catheter and indwelling device-associated infections. The ability of CoNS, including the archetypal species Staphylococcus epidermidis, to cause disease depends on their ability to adhere to polymer surfaces where they form thick, multilayered, cellular agglomerations known as biofilm. The principal components of S. epidermidis biofilm include poly-β-1,6-N-acetyl-D-glucosamine (PNAG, also known as PIA for polysaccharide intercellular adhesion), synthesized by the products of the ica genes, and surface proteins such as the accumulation associated protein Aap. Biofilm contributes to persistence by limiting efficacy of antibiotics and host immune responses. Bloodstream and urinary tract infections ranked as 2nd and 3rd causes respectively of healthcare-associated deaths in the US in 2002. More than 5 million central venous catheters are inserted annually in the U.S. and of the more than 200,000 healthcare-acquired bloodstream infections that occur annually, most are due to central venous catheter. These infections lead to increased morbidity, mortality, lengths of hospitalization, and total healthcare costs.
Many drugs and compounds have been tested as biofilm inhibitors and some are used to coat catheters (e.g. silver, minocycline, rifampin, platinum, nitrofurantoin, chlorhexidine, and sulfadiazine). Several randomized trials have shown benefits of using antibiotic(s)-impregnated catheters in hospitalized patients to reduce colonization and catheter-related bloodstream infections (CRBSI) and include: chlorhexidine-silver sulfadiazine impregnated catheters compared with non impregnated catheters; rifampin-minocycline coated catheters compared with non-coated catheters; and rifampin-minocycline impregnated catheters compared with chlorhexidine-silver sulfadiazine catheters.
Nitazoxanide (NTZ) is a 5-nitrothiazole therapeutic that is used to treat a wide variety of parasitic and anaerobic bacterial infections and is FDA approved for treatment of Cryptosporidium parvum and Giardia intestinalis infections in adults and children. The drug also shows efficacy against Clostridium difficile infections. Mechanistic studies have shown that NTZ is a potent inhibitor of pyruvate: ferredoxin oxidoreductase and therefore is active against all organisms (anaerobic bacteria and parasites) expressing this enzyme. Mechanistic studies revealed that the anionic form of the drug is biologically active and a proton abstraction mechanism has been proposed. Such a generic mechanism might account for the wide range of biological targets reported for this drug.
Mechanistic studies have shown that NTZ is a potent inhibitor of PFOR by interfering with the function of the thiamine pyrophosphate cofactor. The anion form of the drug abstracts a proton from the activated TPP complex and thereby blocks catalysis of pyruvate to acetyl CoA and CO2. The protonated form of NTZ is biologically inactive. Staphylococcal species utilize pyruvate dehydrogenase and not PFOR to catalyze the oxidative decarboxylation of pyruvate. However, the chemical reactivity of NTZ might not be limited to the PFOR target as the drug has been shown to inhibit nitroreductases, protein disulfide bond isomerases and other targets.
Enteroaggregative Escherichia coli (EAEC) strains have emerged as a common cause of persistent diarrhea and malnutrition among children and HIV-infected persons. During infection, EAEC typically adheres to the intestinal mucosa via fimbrial adhesins that results in a characteristic aggregative pattern.
Infectious diarrheal diseases are the second highest global cause of morbidity and mortality, and repeated or prolonged episodes of diarrhea can stunt the growth of infected children and impair cognition. The World Health Organization has estimated that stunting affects approximately 147 million children in the developing world, where every child less than five years old suffers an average of three diarrheal episodes per year. Due to the morbidity burden of diarrheal disease, especially during early childhood, more effective therapies are expected to save many disability-adjusted life years.
EAEC, first identified and described as a diarrheagenic E. coli in 1987, has emerged as a leading cause of acute and persistent (≧14 days) diarrhea among children, AIDS patients, and international travelers in developing and industrialized countries. Around the world, EAEC accounts for 8-32% of acute diarrhea cases among infants and children and 20-30% of persistent diarrhea cases. Individuals most often contract infection via the fecal-oral route by consuming contaminated food and water or by practicing poor hygiene.
The clinical presentation of EAEC infection often consists of watery diarrhea, at times with passage of blood and mucus, but some infections are asymptomatic. This phenomenon is likely due to differences in both host susceptibility and strain heterogeneity. Patients often experience intestinal inflammation marked by elevated levels of fecal lactoferrin, and EAEC infection may perpetuate childhood malnutrition.
EAEC pathogenesis is complex and not fully understood, in large part due to the heterogeneity of strains. Generally, EAEC pathogenesis involves three stages: 1) adherence to the intestinal mucosa, mediated by aggregative adherence fimbriae (AAF); 2) biofilm formation on the surface of host enterocytes; and 3) release of EAEC toxins, elicitation of an inflammatory response, intestinal secretion, and mucosal toxicity, which results in microvillus vesiculation and epithelial cell extrusion.
Several virulence factors have been implicated in mucosal adherence and biofilm formation. Most important and best-studied is the master transcriptional regulator AggR, whose gene is located on a 60-65 MDa pAA plasmid present in many, but not all, strains of EAEC. AggR is activated in response to environmental cues such as low sodium, oxygen, pH and nutrients and controls the expression of several plasmid-encoded genes involved in fimbrial biogenesis, notably aafA (GenBank accession no. AF012835), which encodes for a major structural subunit of the AAF/II variant, expressed by the pathogenic strain 042. AAF/II fimbriae have been described as 5 nm in diameter, arranged in semirigid, filamentous bundles (7), and are thought to mediate adherence to the colonic mucosa and to polystyrene and glass surfaces. AggR also controls expression of other fimbrial genes (e.g., AAF/I, AAF/III) that are antigenically different, some of which can agglutinate erythrocytes or have other non-biofilm producing phenotypes.
Following adherence, EAEC produces a mucosal biofilm that promotes colonization and resists penetration of antimicrobials. The biofilm of strain 042 consists of thick aggregates of bacteria interspersed with void spaces, similar to other bacterial biofilms and characteristic of the heavy biofilm that forms over the epithelium during infection. Biofilm formation is mediated by AAF fimbriae whose expression involves two chromosomal genes, fis and yafK, which are activated by AggR. However, the genetic markers that characterize biofilm-producing strains and the clinical relevance of biofilm formation during infection remain unclear.
There is a long felt need in the art for new and improved antimicrobial agents. The present invention satisfies these needs.